Effects of changes in inspired oxygen fraction on urinary oxygen tension measurements

Background Continuous measurement of urinary PO2 (PuO2) is being applied to indirectly monitor renal medullary PO2. However, when applied to critically ill patients with shock, its measurement may be affected by changes in FiO2 and PaO2 and potential associated O2 diffusion between urine and ureteric or bladder tissue. We aimed to investigate PuO2 measurements in septic shock patients with a fiberoptic luminescence optode inserted into the urinary catheter lumen in relation to episodes of FiO2 change. We also evaluated medullary and urinary oxygen tension values in Merino ewes at two different FiO2 levels. Results In 10 human patients, there were 32 FiO2 decreases and 31 increases in FiO2. Median pre-decrease FiO2 was 0.36 [0.30, 0.39] and median post-decrease FiO2 was 0.30 [0.23, 0.30], p = 0.006. PaO2 levels decreased from 83 mmHg [77, 94] to 72 [62, 80] mmHg, p = 0.009. However, PuO2 was 23.2 mmHg [20.5, 29.0] before and 24.2 mmHg [20.6, 26.3] after the intervention (p = 0.56). The median pre-increase FiO2 was 0.30 [0.21, 0.30] and median post-increase FiO2 was 0.35 [0.30, 0.40], p = 0.008. PaO2 levels increased from 64 mmHg [58, 72 mmHg] to 71 mmHg [70, 100], p = 0.04. However, PuO2 was 25.0 mmHg [IQR: 20.7, 26.8] before and 24.3 mmHg [IQR: 20.7, 26.3] after the intervention (p = 0.65). A mixed linear regression model showed a weak correlation between the variation in PaO2 and the variation in PuO2 values. In 9 Merino ewes, when comparing oxygen tension levels between FiO2 of 0.21 and 0.40, medullary values did not differ (25.1 ± 13.4 mmHg vs. 27.9 ± 15.4 mmHg, respectively, p = 0.6766) and this was similar to urinary oxygen values (27.1 ± 6.17 mmHg vs. 29.7 ± 4.41 mmHg, respectively, p = 0.3192). Conclusions Changes in FiO2 and PaO2 within the context of usual care did not affect PuO2. Our findings were supported by experimental data and suggest that PuO2 can be used as biomarker of medullary oxygenation irrespective of FiO2. Supplementary Information The online version contains supplementary material available at 10.1186/s40635-022-00479-y.

Background Critically ill patients with sepsis develop acute kidney injury (AKI) in 20 to 50% of cases [1]. Despite its clinical importance, available methods for early detection of kidney damage have major limitations. One of the mechanisms implicated in the pathophysiology of this condition is hypoxia of renal tissue, particularly in the renal medulla [2]. Selective hypoxia in the renal medulla was observed in an ovine model of sepsis, despite the presence of global renal hyperemia [3]. In clinical practice, it is not feasible to measure renal medullary tissue oxygen tension in the intensive care unit (ICU). Nevertheless, it is possible to measure oxygen tension of bladder urine (PuO 2 ) by the introduction of a fiber-optic probe in the bladder catheter [4,5]. A number of experimental investigations have been performed to evaluate this technique and have documented a robust correlation between urinary PO 2 and medullary PO 2 [6].
One of the caveats for the understanding of relationship between urinary and medullary PO 2 is that it may be cofounded by several factors. A concern arising from experimental and computational models is that systemic arterial oxygen tension (PaO 2 ) may affect ureteric and bladder wall oxygenation which, in turn, may influence PuO 2 [7][8][9][10]. Experimental observations in anesthetized rabbits suggested that oxygen exchange within the urinary tract is slow and is unlikely to be a major confounder of the relationship between renal medullary tissue PO 2 and PuO 2 [7]. Nevertheless, the potential for such confounding in human sepsis, where PuO 2 measurement might be applied to guide management of risk of AKI and therapeutic interventions, remains unknown. Therefore, to assess whether systemic oxygenation has a potentially confounding impact on urinary oxygenation, we aimed to evaluate PuO 2 in critically ill patients with sepsis during the periods before and after changes in fractional inspired oxygen (FiO 2 ) instituted to manage PaO 2 within clinically acceptable levels. Also, to support our clinical findings, we investigated the medullary and urinary oxygen tension levels in a sheep experiment within a similar range of FiO 2 variation.

Study design
We performed a prospective observational cohort study in the ICU of a tertiary care hospital located in Melbourne, Australia, from January 2017 to March 2018. The protocol was approved by the Human Research Ethics Committee of the Austin Hospital (HREC/16/Austin/26). Written informed consent was obtained from all participants or their legal representatives.

Participants
A convenience sample of adult patients (18 years old or older) with suspected or confirmed septic shock was enrolled in the study. We excluded patients who were anuric, on chronic dialysis, pregnant, or who were kidney transplant recipients.

Measurement of PuO2
For each patient, a fiberoptic luminescence optode (NX-LAS-1/O/E-5 m, Oxford Optronix, Abingdon, UK) was inserted into the lumen of the urinary catheter via a sterile procedure, as described in detail previously [11]. In brief, the sensing tip of the probe was advanced to the distal tip of the catheter so that the probe was placed inside the bladder. The fiberoptic probe was connected to a luminescence lifetime oximeter (Oxylite Pro, Oxford Optronix, UK) interfaced with a laptop computer running LabChart software (Version 8, ADInstruments, Bella Vista, NSW, Australia). PuO 2 was recorded every minute from the time of the probe insertion until the removal of the urinary catheter by the treating medical team or at ICU discharge, whichever occurred first.

FiO2 settings and measurement of PaO2 and PuO2
FiO 2 was modified at the discretion of bedside clinicians to maintain a peripheral oxygen saturation level greater than 90%. Episodes of FiO 2 change were documented in the laptop computer software by the bedside nurse at the moment the intervention took place and were verified against observation charts. To describe arterial oxygen levels, we identified arterial blood gases (ABG) that were collected before and after FiO 2 modification (Fig. 1). To make before and after periods distinctive, we restricted the analysis to ABG samples that were obtained within a time difference from FiO 2 change of 30 min or more. For each PaO 2 measurement, we obtained the mean value of 30 PuO 2 measurements centerd around the exact time when the ABG was collected (15 min before and 15 min after the blood was drawn). Blood gas analysis was performed with an ABL800 FLEX blood gas machine (Radiometer, Copenhagen, Denmark). No specific method was used to ascertain the ABG stability for the purposes of the study. However, the unit where the study was conducted is a world-class intensive care with a 1:1 nurse-to-patient ratio. The ABG analyzer is located inside the unit and the team is trained to obtain reliable measurements according to the institutional protocol.

Data extraction
Aside from the abovementioned variables, we collected information on age, gender, baseline creatinine, infection source, comorbidities and ICU severity scores. We also recorded data on duration of mechanical ventilation, number of hours in mandatory and spontaneous modes, mechanical ventilation parameters, end-tidal partial pressure of carbon dioxide (EtCO 2 ), ICU and hospital length of stay, and hospital mortality.

Animal preparation
We obtained data from 9 healthy adult Merino ewes included in an experimental study of sheep undergoing aseptic surgical procedures under general anesthesia [18]. The study was approved by the Animal Ethics Committee of the Florey Institute of Neuroscience and Mental Health under guidelines laid down by the National Health and Medical Research Council of Australia. A similar fiberoptic luminescence optode (Oxford Optronix, Abingdon, UK) used in human patients was inserted into the lumen of the urinary catheter and surgically inserted into the renal medulla. The tissue oxygen tension was continuously recorded at 100 Hz on a computer using a CED micro 1401 interface with Spike 2 software (Cambridge Electronic Design, Cambridge, United Kingdom).

Experimental protocol for the variation of FiO2
The protocol had 4 components of 20-min duration: a 10-min period to allow oxygen levels to stabilize followed by a 10-min experimental period. Our primary experimental stabilization criteria included renal medullary PO 2 and urinary PO 2 . This timing was determined by assessing the medullary and urinary PO 2 values over time. A block randomization was used to set FiO 2 at 0.21, 0.40, 0.60 and 1.0. The total gas flow on the mechanical ventilator was maintained at a constant rate of 1.5 L/min, whilst the ratio of the individual oxygen-to-air gas volumes was altered to achieve the target FiO 2 . For the current analysis, we obtained data from the periods when FiO 2 levels were 0.21 and 0.40.

Statistical analyses
Continuous variables are reported as median (quartile 1, quartile 3) and categorical variables are reported as number (%). An aggregate measure was calculated per patient and the paired-sample Wilcoxon test was used to compare median values between the two time periods (before and after FiO 2 change). Proportions were compared using Fisher's exact test. To account for multiple episodes of FiO 2 change per patient, we performed a mixed linear regression model to assess the relationship between the variation of PaO 2 (ΔPaO 2 ) and the variation of PuO 2 (ΔPuO 2 ).
In the sheep experiment, values for each FiO 2 setting were calculated and a comparison between groups was performed by using Kruskal-Wallis test.
Statistical analysis was performed using R version 4.0.5. Two-tailed p ≤ 0.05 was considered statistically significant.

Human septic patients
We studied 10 patients, whose clinical characteristics are reported in Table 1. During the study period, patients were mechanically ventilated for 733 h (88.9% of total duration), of which 459 h were in spontaneous mode (55.6%). The mechanical ventilation parameters during the study period are described in Table 2. Arterial blood gases were obtained in 233 occasions. Across the cohort of 10 patients there was a weak but statistically significant positive association between PaO 2 and PuO 2 (Fig. 2, r 2 = 0.022, p = 0.004). This relationship was plotted for each patient (Additional file 1: Fig. S1).
We  Fig. 3). Nevertheless, PuO 2 did not vary significantly across the two time points, being 23.2 [20.5, 29.0] mmHg before the intervention and 24. Other laboratory parameters measured before and after FiO 2 change were similar between the two time points (Table 3). The urine output before FiO 2 change was 63.

Experimental study
In the sheep experiment, we evaluated urinary and medullary tissue oxygen measurements in four FiO 2 levels: 0.21, 0.40, 0.60 and 1.00 (Table 4)

Key findings
We conducted an observational study in human septic patients to determine whether PuO 2 is affected by changes in systemic oxygenation during routine care of patients with septic shock. As expected, changes of FiO 2 resulted in significant changes in PaO 2 . However, we found no significant differences between PuO 2 measured before and after the interventions occurred. We supported our clinical findings with data from an experimental sheep experiment showing that medullary and urinary oxygen tension measurements did not differ within a similar range of FiO 2 variation.

Relationship to previous studies
To our knowledge, there have been no previous investigations of the relationship between systemic and urinary oxygenation in human patients with septic shock. Ngo et al. addressed this issue in a group of patients undergoing cardiac surgery, finding no significant relationship between PaO 2 and PuO 2 during cardiopulmonary bypass [6]. Importantly, however, these observations were obtained in a unique physiological state with non-pulsatile flow, an extracorporeal circuit and mild hypothermia. Our current observations are likely more generally applicable to patients in a critical care setting. They are also very consistent with the outcomes of a retrospective analysis of three 3 studies involving a total of 28 adult Merino ewes during experimental sepsis [4,12,13], in which only a weak linear relationship was found between PaO 2 and PuO 2 , accounting for ≤ 6% of the variation of PuO 2 [6].
The absence of detectable changes in PuO 2 in response to modest but clinically significant changes in FiO 2 and thus PaO 2 indicate that renal medullary tissue PO 2 was not markedly affected by these clinical maneuvers. Experimental evidence supports the concept that extreme variations in FiO 2 and/or PaO 2 lead to corresponding changes in the oxygen tension of renal tissue. For example, in anesthetized rats a reduction in FiO 2 from 1.0 to 0.1 resulted in a decline in cortical and medullary microvascular PO 2 as assessed by dual-wavelength phosphorimetry [14]. Likewise, studies using fluorescence optodes in anesthetized rabbits demonstrated variations in both cortical and medullary tissue PO 2 with variations in FiO 2 [7,[15][16][17]. These findings provide support to our experimental findings that greater FiO 2 variation is associated with greater PuO 2 response, in particular at 0.60 and 1.00 FiO 2 . Our ovine study sample derived from a larger experiment comprising 18 healthy sheep undergoing abdominal surgery under total intravenous or volatile anesthesia. In this study, increasing FiO 2 from 0.21 to 1.00 increased cortical and medullary tissue PO 2 [18]. However, it is also well-established that renal medullary tissue PO 2 is less responsive to changes in PaO 2 than the renal cortical tissue PO 2 [16,18,19]. This appears to be a consequence of counter-current diffusive shunting of oxygen between descending and ascending vasa recta, which acts to reduce delivery of oxygen to renal medullary tissue [20]. Consequently, small but physiologically (and clinically) significant changes in FiO 2 and/or PaO 2 may not appreciably alter renal medullary tissue PO 2 . In support of this concept, no appreciable difference was observed in medullary tissue PO 2 in our group's preceding experiment when FiO 2 was varied from 0.4 to 0.6 [18]. Similarly, in anesthetized rats outer medullary microvascular PO 2 did not vary significantly when FiO 2 was varied from 0.21 and 0.30 [21]. We cannot directly measure renal medullary tissue PO 2 in patients and can only draw indirect inferences from measurement of PuO 2 and consideration of available experimental evidence. However, the most parsimonious interpretation of our current findings is that modest changes in FiO 2 and thus PaO 2 neither markedly alter renal medullary tissue PO 2 in patients with sepsis nor confounded the relationship between medullary tissue PO 2 and PuO 2 .

Study implications
Our findings suggest that commonly performed adjustments to FiO 2 settings in patients with sepsis do not result in significant changes in PuO 2 . In consonance of these findings, we observed that variations of FiO 2 between 0.21 and 0.40 did not alter either medullary or urinary oxygen tension measurements in a sheep experiment. Thus, variations of systemic oxygenation seem unlikely to confound or affect the utility of urinary oxygenation as a biomarker for risk of AKI. Nevertheless, at higher FiO 2 (0.60 and 1.00), significantly increased PuO 2 values were obtained. One possible explanation is that in our septic patients, the FiO 2 gap was far smaller in comparison to the experimental study. Also, one could argue that a type 2 error was present in the observational study which may have been controlled for during the experimental protocol.
Additional investigation is needed to explore whether the lack of PuO 2 variation in face of PaO 2 changes derives from the presence of confounding factors affecting medullary oxygen values. Also, further studies in critically ill patients are needed to elucidate whether sustained differences in oxygen exposure [22,23] influence renal related outcomes. Thus, changes in PaO 2 , as a consequence of altered FiO 2 in routine care of patients with septic shock, is unlikely to be a major confounder of the relationship between renal medullary tissue PO 2 and PuO 2 . In the current study, PaO 2 was used as a measure of systemic oxygenation because it reflects the balance between oxygen delivery and consumption. Had SpO 2 been used, the accuracy would have been affected by peripheral tissue perfusion, use of vasoactive agents and altered cardiac output. Finally, continuous measurement of PuO 2 might be useful for monitoring the impact on renal medullary oxygenation.

Strengths and limitations
We evaluated systemic and urinary oxygenation in human septic patients and assisted our proposition with experimental data. Our findings are consistent with previous observations in sepsis [6] and provide additional evidence that the relationship between renal medullary tissue PO 2 and PuO 2 is unlikely to be confounded by changes in FiO 2 or PaO 2 in the range commonly encountered in the ICU. As such, they provide further support for the use of PuO 2 as a clinical surrogate of renal medullary PO 2 . Our study has several limitations. First, the clinical component was an observation designed to assess the effects on PuO 2 where the observed intervention (change of FiO 2 ) was not protocolized. Moreover, controlling for variables such as creatinine or urine output was not feasible due to technical limitations and the limited number of patients. However, we aimed to undertake an exploratory analysis to generate a preliminary hypothesis to guide advanced studies. Moreover, we added data from a sheep experiment where FiO 2 variation was protocolized. Also, due to the lower number of measurements in the experimental study, greater heterogeneity was observed. Second, the inclusion of septic patients in our clinical study did not occur in the early stage of resuscitation. On the other hand, the instances of FiO 2 change we captured took place in a stable state with lower propensity for PuO 2 to be affected by additional confounding effects of interventions intended to optimize oxygen delivery to the tissues. Furthermore, changes in FiO 2 performed under stable conditions might have reduced the likelihood of reverse causation or provided mitigation of any potential effect of other interventions. Third, the observational nature of the study may have led to confounding by indication. For instance, the reasons motivating the clinician to change FiO 2 settings could have affected the relationship between FiO 2 and PuO 2 . However, a larger degree of FiO 2 change would be expected if optimization measures capable of affecting such relationship were in place. Our patients were enrolled in the stabilization phase of sepsis, a time when, in general, only limited interventions are performed to achieve physiologic parameters aiming to prevent organ dysfunction. Finally, we addressed only the variation of systemic oxygenation within the normoxemic range. However, such a normoxemic range is typical in the care of patients in the ICU.

Conclusions
Changes in FiO 2 and PaO 2 within the context of usual care did not appreciably affect PuO 2 . Our findings suggest that, within the values reported, PuO 2 measured in a clinical and experimental setting is not confounded by changes in inspired oxygen fraction or arterial oxygen tension and that PuO2 can be used as biomarker of medullary oxygenation irrespective of FiO2.

ABG
Arterial blood gas test AKI Acute kidney injury EtCO 2 End-tidal partial pressure of carbon dioxide FiO 2 Fraction of inspired oxygen. ICU Intensive Care Unit PaO 2 Arterial oxygen tension PuO 2 Urinary oxygen tension